Capillary

Electrophoresis
 Electrophoresis—An Overview
• Definition: The differential
movement for migration of ions by
attraction or repulsion in an
electric field.
– Separation of components of a
mixture using an electric field
• v=Eq/f
– v = velocity of molecule
– E = electric field
– q = net charge of molecule
– f = friction coefficient
Electrophoresis- overview cont.
• Can determine the size, shape, and
charge of a molecule
• Different forms of electrophoresis
are used for each of these factors
independently or in combination.
 Types of Electrophoresis
• Capillary
• Native Polyacrylimide Gel
Electrophoresis (PAGE)
• SDS-PAGE
• Slab
• Paper
Capillary Electrophoresis
Apparatus
Capillary Electrophoresis – The
Basics
• Electrophoresis in a buffer filled,
narrow-bore capillaries
• Each capillary is about 25-100 μm in
internal diameter
• When a voltage is applied to the
solution, the molecules move through
the solution towards the electrode of
opposite charge
• Depending on the charge, the
molecules move through at different
speeds
– Separation is achieved
Basics cont.
• A photocathode is
then used to
measure the
absorbencies of the
molecules as they
pass through the
solution
• The absorbencies
are analyzed by a
computer and they
are represented
graphically
Ooooo!
It
MOVE
S
 Equipment
• Capillary tube
• Varied length but
normally 25-50 cm
• Small bore and
thickness of the silica
play a role
– Using a smaller internal
diameter and thicker
walls help prevent Joule
Heating, heating due to
voltage
 Equipment

• Because ions are in the bulk solution are about to travel

through the capillary without interference from the
capillary itself, there is no dramatic drop in potential
within the capillary
• No meniscus is made
 Yowzah!!
Equipment !

• Detector
– UV/Visible
absorption
– Fluorescence
– Radiometric (for
radioactive
substances)
– Mass Spec.
 Applications
• Analysis of • Advantages
carbohydrates • Fast
• Small Sample
• Analysis of
• Relatively
inorganic inexpensive
anions/metal ions • Automated
• DNA profiling • Disadvantages
• Cannot identify
• Protein
neutral species
identification • Joule Heating
• Cannot discern
shape
 Resources
• Altria, Kevin. http://www.ceandcec.com
• Hardy, James K. University of Akron.
http://ull.chemistry.uakron.edu/chemsep/electrophoresis/
• University of Virginia Chemistry Web page.
http://www.virginia.edu/chem/people/
• http://www.rsc.org/pdf/books/capelectrosc.pdf
• Landers, James. Electrophoresis SDS-PAGE.
http://toolkit.itc.virginia.edu/cgi-local/tk/UVa_CLAS_2002_Fall_C